The precise pattern of input and output connections in the brain is achieved through tightly regulated developmental processes whose impairment have profound effects on brain function, and are linked to neurodevelopmental defects ranging from mental retardation and autism spectrum disorders. Upon reaching their targets, axons branch extensively and the size of their terminal arborization defines their synaptic output. In many axons including cortical projections, axon branching is regulated by both activity-independent and activity-dependent mechanisms. However, the molecular mechanisms controlling axon branching of mammalian neurons in vivo are still poorly understood. We recently identified a new kinase pathway defined by LKB1 and one of its 14 direct downstream kinases called NUAK1 as critical regulators of cortical axon branching in vivo. We found that the LKB1-NUAK1 kinase pathway regulates axon branching through promoting presynaptic mitochondrial capture. These results raise one central unresolved question: how do presynaptic mitochondria regulate axon branching? More generally, little is known about the function of presynaptic mitochondria during axon development beyond their function as ATP provider. We have consolidated exciting new results showing that presynaptic mitochondria play a critical role in presynaptic calcium homeostasis through the Mitochondrial Calcium Uniporter (MCU). Therefore, we have emphasized the revised proposal on studying the function of LKB1 in regulating (1) presynaptic mitochondrial capture and/or (2) the Ca2+ clearance capacity of presynaptic mitochondria. We propose that there are two phenotypes in LKB1 and maybe in NUAK1-null axons that both contribute to the reduction in axon branching: (1) the failure of most nascent presynaptic sites to capture mitochondria during early development (which results in lack of mitochondria-dependent calcium clearance) and (2) at the few presynaptic sites were mitochondria are found in LKB1-null axons (25% instead of 70% in wild-type axons), these mitochondria have reduced MCU expression which correlates with reduced mitochondria-dependent presynaptic calcium clearance. We present preliminary results showing that this increased presynaptic calcium accumulation has drastic consequence on presynaptic release properties in LKB1-null axons. This grant is focusing on further exploring the relationship between LKB1 signaling, mitochondria function in presynaptic calcium homeostasis, presynaptic release properties and axon branching. The proposed experiments will provide important new insights into the molecular and cellular mechanisms underlying the development of cortical connectivity, a critical step for the emergence of cognitive functions.

Public Health Relevance

This proposal is aimed at determining the role of LKB1-NUAK1 kinases during brain development and in particular to study the function of this new kinase pathway in the development of cortical connectivity. Loss-of-function mutations in NUAK1 and other LKB1-dependent kinases have recently been identified in patients diagnosed with Autism Spectrum Disorder, therefore our results will elucidate how genetic mutation of NUAK1 contributes to abnormal brain development in this socially-devastating neurodevelopmental disorder.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Neurodifferentiation, Plasticity, and Regeneration Study Section (NDPR)
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Riddle, Robert D
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Columbia University (N.Y.)
Schools of Medicine
New York
United States
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Toyama, Erin Quan; Herzig, Sébastien; Courchet, Julien et al. (2016) Metabolism. AMP-activated protein kinase mediates mitochondrial fission in response to energy stress. Science 351:275-81
Shimojo, Masafumi; Courchet, Julien; Pieraut, Simon et al. (2015) SNAREs Controlling Vesicular Release of BDNF and Development of Callosal Axons. Cell Rep 11:1054-66
Hsia, Hung-En; Kumar, Rohit; Luca, Rossella et al. (2014) Ubiquitin E3 ligase Nedd4-1 acts as a downstream target of PI3K/PTEN-mTORC1 signaling to promote neurite growth. Proc Natl Acad Sci U S A 111:13205-10
Courchet, Julien; Lewis Jr, Tommy L; Lee, Sohyon et al. (2013) Terminal axon branching is regulated by the LKB1-NUAK1 kinase pathway via presynaptic mitochondrial capture. Cell 153:1510-25